Total synthesis of pinellic acid, a potent oral adjuvant for nasal influenza vaccine. Determination of the relative and absolute configuration

Article abstract of DOI:10.1016/S0040-4039(01)02348-6

Pinellic acid (1), isolated from a medicinal plant Pinelliae tuber, has potent adjuvant activity. The absolute configuration of pinellic acid was expected by derivatization of this compound and CD exciton chirality method. A convergent synthetic route to pinellic acid has been developed via regioselective asymmetric dihydroxylation and stereoselective reduction. The absolute configuration of pinellic acid was determined to be 9S,12S,13S, by comparing with the spectra data of natural and synthetic compounds.

Full text of DOI:10.1016/S0040-4039(01)02348-6

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 43 (2002) 1265–1268
Total synthesis of pinellic acid, a potent oral adjuvant for nasal
influenza vaccine. Determination of the relative and absolute
configuration
Toshiaki Sunazuka,b,e Tatsuya Shirahata,b Kiminari Yoshida,b Daisuke Yamamoto,b
Yoshihiro Harigaya,b Takayuki Nagai,c,d Hiroaki Kiyohara,c,d Haruki Yamada,c,d Isao Kuwajimad,e
a,d,
(
and Satoshi Omura
*
aThe Kitasato Institute, Shirokane, Minatoku, Tokyo 108–8641, Japan
bSchool of Pharmaceutical Sciences, Kitasato University, Shirokane, Minatoku, Tokyo 108–8641, Japan
cOriental Medicine Research Center, the Kitasato Institute, Shirokane, Minatoku, Tokyo 108–8641, Japan
dKitasato Institute for Life Sciences, Kitasato University, Shirokane, Minatoku, Tokyo 108–8641, Japan
eCREST, The Japan Science and Technology Corporation (JST), Shirokane, Minatoku, Tokyo 108–8641, Japan
Received 12 November 2001; revised 3 December 2001; accepted 7 December 2001
Abstract—Pinellic acid (1), isolated from a medicinal plant Pinelliae tuber, has potent adjuvant activity. The absolute configura-
tion of pinellic acid was expected by derivatization of this compound and CD exciton chirality method. A convergent synthetic
route to pinellic acid has been developed via regioselective asymmetric dihydroxylation and stereoselective reduction. The absolute
configuration of pinellic acid was determined to be 9S,12S,13S, by comparing with the spectra data of natural and synthetic
compounds. © 2002 Elsevier Science Ltd. All rights reserved.
Influenza virus infection is epidemic, and sometimes
critical for patients with respiratory diseases and aged
persons with respiratory diseases. Influenza vaccine is
useful as prophylaxis of influenza virus infection.
Intranasal inoculation of influenza vaccine has been
tried to increase its safety and to prevent antigenic
variation of influenza viruses. Nonetheless, it has gener-
ally been considered that intranasal inoculation of the
vaccine alone cannot readily induce high levels of anti-
bodies. Therefore, the development of effective adju-
vants for nasal influenza vaccine is necessary to
enhance the potency of the vaccine.
Pinelliae tuber as an active compound of the adjuvant
activity4 (Fig. 1).
Although 1 is an effective oral adjuvant for nasal
influenza vaccine, Pinelliae tuber contains a small
amount of
1 and the stereochemistry remained
unknown. Although 10E-9,12,13-trihydroxyoctade-
cenoic acid as 1 has been isolated from natural sources,
we were not able to determine its relative and absolute
stereochemistry because of lack of detailed reports
regarding its stereochemistry.5 Herein, we report the
enantioselective total synthesis and assignment of the
absolute stereochemistry of 1.
In our program to discover effective adjuvants, Kampo
medicine, ‘Sho-seiryu-to’ was found to possess potent
adjuvant activity by oral administration on nasal influ-
enza infection and nasal influenza vaccination.1–3 Fur-
thermore, our research has made it clear that the
activity of ‘Sho-seiryu-to is expressed by ingredients
from Pinelliae tuber, one of the component herbs of
‘Sho-seiryu-to’. Pinellic acid 1 has been isolated from
First, to determinate the relative configuration of 12,13-
diol, esterification of 1 followed by dimethylacetaliza-
tion gave acetonide 2 (Scheme 1). The coupling
Keywords: adjuvant; 10E-9,12,13-trihydroxyoctadecenoic acid; asym-
metric dihydroxylation; BINAL-H.
* Corresponding author. Tel.: +81-3-3444-6380; fax: +81-3-3444-6380;
e-mail: omura-s@kitasato.or.jp
Figure 1. Structure of pinellic acid 1.
0040-4039/02/$ - see front matter © 2002 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(01)02348-6

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T. Sunazuka et al. / Tetrahedron Letters 43 (2002) 1265–1268
Scheme 1. Derivatization of 1. Reagents and conditions: (a) (1) TMSCHN2, benzene:MeOH (10:1), rt, 2.5 h, (2) 2,2-
dimethoxypropane, PPTS, CH2Cl2, 60°C, 48 h, (100% from 1); (b) p-Br-BzCl, DMAP, pyridine, (68%).
1
constant (J12,13=8.0 Hz) in the H NMR spectrum of 2
and NOE analysis indicates syn configuration at 12,13-
diol (Fig. 2).
shows a positive Cotton effect [umax (Dm): 244.8 (+6.97),
220.8 (+2.13), 209.1 (+5.97) (MeOH)], which suggested
9S configuration (Fig. 3).7
On the other hand, to establish the absolute configura-
tion of C9 by CD exciton,6 the corresponding p-bromo-
benzoate 3 was prepared. The coupling constant
Based on these results the absolute configuration of 1
was assumed as 4 (9S,12S,13S) or 5 (9S,12R,13R) (Fig.
4). Accordingly, we tried to establish a convergent
synthetic route to 4 and 5.
1
between 9H and 10H in the H NMR spectrum of 3
was 7.0 Hz, indicating antiperiplanar conformation of
these two protons. Moreover, the CD spectrum of 3
The strategic disconnection is outlined in Fig. 5. The
main problem should be the construction of stereo-
chemistry of three hydroxy groups. We planned to
construct the syn diol at C12-13 by regioselective asym-
metric dihydroxylation.8 The allylic alcohol at C9 could
be prepared from enone by stereoselective reduction.
The synthesis of C18 skeleton 11 is shown in Scheme 2.
The esterification of suberic acid mono methylester 6
with (Boc)2O, DMAP in t-BuOH furnished tert-
butylester 7. The diester 7 was converted to iodide 8 in
good yield via three steps: (1) hydrolysis of methyl
ester, (2) reduction of carboxylic acid9 and (3) iodina-
tion of the prepared primary alcohol. The dithiane
coupling10 of 10 derived from commercially available 9
with 8 gave diene 11 in high yield. We next introduced
hydroxy groups to the C18 skeleton (Scheme 3).
Figure 2. NOE experiment of 2.
The regioselective asymmetric dihydroxylation of 11 by
using modified Sharpless ligand (DHQ(PHAL)DHQ·
Me+I−)8 furnished 12,13 syn diol (−)-12 which has
12S,13S in 75% yield with 95% e.e. followed by protec-
tion of diol with TBSOTf.12 Deprotection of dithioac-
etal (+)-13 provided enone (+)-14. The reduction of
enone (+)-14 was reported.13 We found (S)-BINAL-H14
was the best reagent for stereoselective reduction of
(+)-14 to furnish 9S alcohol11 (diastereoselectivity;
>20:1). The desilylation with TBAF gave triol (+)-15 as
a single isomer. The hydrolysis of tert-butyl ester by a
highly concentrated alkaline solution afforded (−)-415
which has 9S,12S,13S configuration. (−)-415 was identi-
1
cal in all respects with natural product 1. [400 MHz H
and 100 MHz 13C NMR, IR, HRMS, optical rotation
{[h]D25 −8.0 (c 0.30, MeOH), natural;4 [h]2D8 −8.1 (c 0.32,
MeOH)}, and oral adjuvant activity].
Figure 3. CD spectrum of 3.
Figure 4. Assumable structure of 1.

T. Sunazuka et al. / Tetrahedron Letters 43 (2002) 1265–1268
1267
On the other hand, for the synthesis (+)-5 according to
the foregoing synthetic route (Scheme 4), asymmetric
dihydroxylation using (DHQD)2PHAL of 11 provided
(+)-12 which has 12R,13R configulation11 in 75% yield
with 92% e.e.. After dihydroxylation, (+)-516 could be
synthesized in the same way for synthesis of (−)-4.
1
(+)-516 was not identical in 400 MHz H and 100 MHz
Figure 5. Synthetic strategy of 4.
Scheme 2. Synthesis of C18 skeleton 11. Reagents and conditions: (a) (Boc)2O, DMAP, t-BuOH, rt, 1 h, (82%), (b) (1) 0.1N NaOH
in THF:MeOH:H2O (3:1:1), rt, 22 h, (2) BH3·THF, THF, 0°C–rt, 24 h, (3) I2, PPh3, imidazole, CH2Cl2, 0°C–rt, 1 h, (77% from
7); (c) 1,3-propanedithiol, BF3·OEt2, CH2Cl2, 0°C–rt, (96%); (d) n-BuLi, THF, −78°C, 1 h, then 8, −78°C, 1 h (85%).
Scheme 3. Synthesis of 4. Reagents and conditions: (a) DHQ(PHAL)DHQ·Me+·I−, K3[Fe(CN)6], K2CO3, K2OsO4·2H2O, methane-
sulfonamide, t-BuOH: H2O (1:1), 0°C, 1 h, (64%, 95% e.e.); (b) TBSOTf, 2,6-lutidine, −78°C, 10 min, (89%); (c) Hg(ClO4)2,
CaCO3, THF:H2O (5:1), rt, 30 min, (83%); (d) (1) (S)-BINAL-H, THF, −78°C, 1 h, (diastereoselectivity; >20:1), (2) TBAF, THF,
70°C, 3 h, (76% from (+)-14); (e) 2.0N KOH in EtOH:H2O (5:1), rt, 46 h, (82%).
Scheme 4. Synthesis of 5. Reagents and conditions: (a) DHQD(PHAL)DHQD, K3[Fe(CN)6], K2CO3, K2OsO4·2H2O, methanesul-
fonamide, t-BuOH:H2O (1:1), 0°C, 1 h, (75%, 92% e.e.), (b) TBSOTf, 2,6-lutidine, −78°C, 10 min, (87%); (c) Hg(ClO4)2, CaCO3,
THF:H2O (5:1), rt, 30 min, (83%); (d) (1) (S)-BINAL-H, THF, −78°C, 1 h, (diastereoselectivity; >20:1); (2) TBAF, THF, 70°C,
3 h, (76% from (−)-14); (e) 2.0N KOH in EtOH:H2O (5:1), rt, 46 h, (76%).

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T. Sunazuka et al. / Tetrahedron Letters 43 (2002) 1265–1268
13C NMR, and optical rotation {[h]2D3 +29.8 (c 0.45,
MeOH)} with natural product 1. The oral adjuvant
activity of synthetic (−)-4 was the same as that of
natural compound 1, while that of (+)-5 was less than 1.
T.; Yoshikawa, K.; Mochida, K.; Hirama, M. Tetra-
hedron Lett. 1999, 40, 8641–8645.
8. Review: Kolb, H. C.; VanNieuwenhze, M. S.; Sharpless,
K. B. Chem. Rev. 1994, 94, 2483–2547.
9. Okuma, K.; Hashimoto, S.; Ono, M.; Shioji, K.; Mat-
suyama, H.; Bestmann, H. J. Tetrahedron. 1998, 54,
4243–4250.
10. Grobel, B. T.; Seebach, D. Synthesis 1977, 357–402.
11. The stereochemistry of all hydroxy groups was deter-
mined by Mosher’s method: Dale, J. A.; Mosher, H. S. J.
Am. Chem. Soc. 1973, 95, 512–519.
In summary, we have elucidated that the relative and
absolute configuration of pinellic acid 1 is 9S,12S,13S.
A synthetic route to pinellic acid was established via
asymmetric dihydroxylation and stereoselective reduc-
tion by (S)-BINAL-H. This route enables preparation
of all of the stereoisomers of 1. Further synthesis of the
isomers and the structure–activity relationship study
are in progress in our laboratory.
12. Torisawa, Y.; Okabe, H.; Ikegami, S. Chem. Lett. 1984,
1555–1556.
13. (a) Suemune, H.; Harabe, T.; Sakai, K. Chem. Pharm.
Bull. 1988, 36, 3632–3637; (b) Ishigami, K.; Motoyoshi,
H.; Kitahara, T. Tetrahedron Lett. 2000, 41, 8897–8901;
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hedron 2001, 41, 3899–3908.
Acknowledgements
14. Noyori, R.; Tomino, I.; Tanimoto, Y.; Nishizawa, M. J.
This work was supported in part by a grant from the
Ministry of Education, Science and Culture of Japan,
and Japan Keirin Association.
Am. Chem. Soc. 1984, 106, 6709–6716.
15. (−)-4: White powder, mp: 104–106°C (MeOH), [h]2D5 −8.0
(c 0.30, MeOH), {natural;4 [h]2D8 −8.1 (c 0.32, MeOH)},
IR (KBr) 3372 (s), 1695 (m), 1637 (m), 1H NMR (400
MHz, CD3OD) l : 5.72 (1H, dd, J=15.5, 5.0 Hz), 5.67
(1H, dd, J=15.5, 5.0 Hz), 4.05 (1H, ddd, J=6.5, 6.0, 5.0
Hz), 3.91 (1H, dd, J=5.5, 5.0 Hz), 3.41 (1H, ddd, J=8.5,
5.5, 2.5 Hz), 2.27 (2H, t, J=7.5 Hz), 1.60 (2H, dt, J=7.5,
7.0 Hz), 1.55–1.50 (2H, m), 1.45–1.25 (16H, m), 0.91 (3H,
t, J=6.3 Hz), 13C NMR (100 MHz, CD3OD) l:177.8,
136.6, 131.1, 76.5, 75.8, 73.0, 38.3, 35.0, 33.6, 33.1, 30.5,
30.4, 30.2, 26.6, 26.5, 26.1, 23.7, 14.4, HR-FABMS m/z:
353.2305 [M+Na], calcd for C18H34NaO5: 353.2304.
16. (+)-5: White powder, mp: 68–71°C (MeOH), [h]2D3 +29.8
(c 0.45, MeOH), IR (KBr) 3430 (s), 1697 (m), 1632 (m),
1H NMR (400 MHz, CD3OD) l : 5.70 (1H, dd, J=15.5,
5.5 Hz), 5.64 (1H, dd, J=15.5, 6.0 Hz), 4.03 (1H, ddd,
6.5, 6.0, 5.5 Hz), 3.87 (1H, dd, J=6.0, 5.5 Hz), 3.40 (1H,
ddd, J=7.0, 5.5, 2.0 Hz), 2.27 (2H, t, J=7.5 Hz), 1.60
(2H, dt, J=7.5, 7.0 Hz), 1.55–1.50 (2H, m), 1.44–1.25
(16H, m), 0.91 (3H, t, J=6.3 Hz), 13C NMR (100 MHz,
CD3OD) l:178.2, 136.7, 131.3, 76.7, 75.7, 73.2, 38.3,
36.0, 33.8, 33.1, 30.5, 30.4, 30.2, 26.5, 26.5, 26.2, 23.7,
14.4, HR-FABMS m/z: 353.2309 [M+Na], calcd for
C18H34NaO5: 353.2304.
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